Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape

Abstract The serine protease inhibitor (SERPIN) plasminogen activator inhibitor-1 (PAI-1) is a key regulator of the fibrinolytic system, inhibiting the serine proteases tissue- and urokinase-type plasminogen activator (tPA and uPA, respectively). Missense variants render PAI-1 non-functional through...

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Autores principales:	Zachary M. Huttinger, Laura M. Haynes, Andrew Yee, Colin A. Kretz, Matthew L. Holding, David R. Siemieniak, Daniel A. Lawrence, David Ginsburg
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f47950a71a2243e4bed673bbf54e9f40
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spelling	oai:doaj.org-article:f47950a71a2243e4bed673bbf54e9f402021-12-02T18:48:09ZDeep mutational scanning of the plasminogen activator inhibitor-1 functional landscape10.1038/s41598-021-97871-72045-2322https://doaj.org/article/f47950a71a2243e4bed673bbf54e9f402021-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-97871-7https://doaj.org/toc/2045-2322Abstract The serine protease inhibitor (SERPIN) plasminogen activator inhibitor-1 (PAI-1) is a key regulator of the fibrinolytic system, inhibiting the serine proteases tissue- and urokinase-type plasminogen activator (tPA and uPA, respectively). Missense variants render PAI-1 non-functional through misfolding, leading to its turnover as a protease substrate, or to a more rapid transition to the latent/inactive state. Deep mutational scanning was performed to evaluate the impact of amino acid sequence variation on PAI-1 inhibition of uPA using an M13 filamentous phage display system. Error prone PCR was used to construct a mutagenized PAI-1 library encompassing ~ 70% of potential single amino acid substitutions. The relative effects of 27% of all possible missense variants on PAI-1 inhibition of uPA were determined using high-throughput DNA sequencing. 826 missense variants demonstrated conserved inhibitory activity while 1137 resulted in loss of PAI-1 inhibitory function. The least evolutionarily conserved regions of PAI-1 were also identified as being the most tolerant of missense mutations. The results of this screen confirm previous low-throughput mutational studies, including those of the reactive center loop. These data provide a powerful resource for explaining structure–function relationships for PAI-1 and for the interpretation of human genomic sequence variants.Zachary M. HuttingerLaura M. HaynesAndrew YeeColin A. KretzMatthew L. HoldingDavid R. SiemieniakDaniel A. LawrenceDavid GinsburgNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-10 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Zachary M. Huttinger Laura M. Haynes Andrew Yee Colin A. Kretz Matthew L. Holding David R. Siemieniak Daniel A. Lawrence David Ginsburg Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
description	Abstract The serine protease inhibitor (SERPIN) plasminogen activator inhibitor-1 (PAI-1) is a key regulator of the fibrinolytic system, inhibiting the serine proteases tissue- and urokinase-type plasminogen activator (tPA and uPA, respectively). Missense variants render PAI-1 non-functional through misfolding, leading to its turnover as a protease substrate, or to a more rapid transition to the latent/inactive state. Deep mutational scanning was performed to evaluate the impact of amino acid sequence variation on PAI-1 inhibition of uPA using an M13 filamentous phage display system. Error prone PCR was used to construct a mutagenized PAI-1 library encompassing ~ 70% of potential single amino acid substitutions. The relative effects of 27% of all possible missense variants on PAI-1 inhibition of uPA were determined using high-throughput DNA sequencing. 826 missense variants demonstrated conserved inhibitory activity while 1137 resulted in loss of PAI-1 inhibitory function. The least evolutionarily conserved regions of PAI-1 were also identified as being the most tolerant of missense mutations. The results of this screen confirm previous low-throughput mutational studies, including those of the reactive center loop. These data provide a powerful resource for explaining structure–function relationships for PAI-1 and for the interpretation of human genomic sequence variants.
format	article
author	Zachary M. Huttinger Laura M. Haynes Andrew Yee Colin A. Kretz Matthew L. Holding David R. Siemieniak Daniel A. Lawrence David Ginsburg
author_facet	Zachary M. Huttinger Laura M. Haynes Andrew Yee Colin A. Kretz Matthew L. Holding David R. Siemieniak Daniel A. Lawrence David Ginsburg
author_sort	Zachary M. Huttinger
title	Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
title_short	Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
title_full	Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
title_fullStr	Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
title_full_unstemmed	Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
title_sort	deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/f47950a71a2243e4bed673bbf54e9f40
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Deep mutational scanning of the plasminogen activator inhibitor-1 functional landscape

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